Abstract Peripheral arterial disease (PAD) is one of the major complications of systemic atherosclerosis where arterial occlusions result in two major clinical manifestations: leg pain with walking/reduced walking ability (a condition termed intermittent claudication, IC) or critical limb ischemia (CLI) with leg pain at rest, ischemic ulcers, and patients that face a >10% risk/year of major amputation. No medical therapy exists to improve perfusion to the ischemic limb in PAD and there remains a clear need to develop new treatments. The central dogma of molecular biology states that DNA makes RNA and RNA makes protein. However, the vast majority of the mammalian genome does not encode proteins. There are two types of non-coding RNAs: micro-RNA (miR) and long non-coding (lncRNA) RNAs. The diversity of miRs across species is far greater than encoded genes and lncRNAs are an order of magnitude more diverse that miRs; this necessitates that strategies study human disease. To that end, this proposal brings together a translational cardiovascular scientist who studies PAD and whose work has included micro-RNAs (Annex), a research scientist who focuses on in-vitro and in-vivo models of PAD (Ganta), a translational scientist who studies skeletal muscle adaptability to injury and has developed an unparalleled biobank of human PAD tissues (McClung), as well as a computational biologist (Keene) to provide bioinformatics analysis of RNA sequencing data. The central hypothesis of this proposal states that condition specific sampling based on insights into the clinical disease condition from well- phenotyped humans, bioinformatics analysis, and disease specific non-clinical testing will unlock non-coding RNA novel human therapeutics. To that end, the Specific Aims are: 1: Identify miRs for therapeutic modulation by determining differential expression in plasma across subjects with intermittent claudication (IC) and critical limb ischemia (CLI) patients and age/sex-matched non-PAD control subjects. Determine whether the ischemic skeletal muscle from the most symptomatic ischemic leg demonstrates elevated expression of the miR. Candidate miR's will be examined through in-vitro studies in PAD relevant conditions, through in-vivo efficacy. 2. Identify lncRNAs for modulation by determining the differential expression of lncRNA in calf muscle from non-PAD and PAD (both IC and CLI) patients. We will first look for lncRNA's that are up-regulated in PAD (IC and CLI) and will then examine whether up- regulation occurs differentially (IC or CLI). In a new cohort, we will examine predictions form an initial list of candidate lncRNAs and will determine whether target up-regulation also occurs in blood. 3) Utilize mouse models of HLI that recapitulate the ?major? differential for the IC vs. CLI designation in the human condition to identify additional candidate lncRNAs that may be conserved from mouse to man and serve as therapeutic candidates.